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Trypanosoma cruzi response to the oxidative stress generated by hydrogen peroxide
Jane K Finzi1, Camila W M Chiavegatto, Karen F Corat
1Departamento de Bioquímica, IB, UNICAMP, Campinas, SP, Brazil.
Abstract:
As an intracellular parasite, Trypanosoma cruzi is exposed to reactive oxygen species. The study of the proteins involved in the hydroperoxide detoxification cascade, tryparedoxin peroxidase included, may lead to the development of a more specific chemotherapy for Chagas'disease. In this work, the involvement of TcCPX in T. cruzi resistance to oxidant-mediated injury was investigated. At low concentrations of hydrogen peroxide cell proliferation was stimulated and parasites increased their resistance to sub-lethal doses of H2O2 (100 microM) if previously treated with a non-toxic concentration of H2O2 (20 microM). Incubation of cells with different H2O2 concentrations induced a dose-dependent increase in TcCPX levels, as detected by Western blotting analysis. The increase in TcCPX levels in the presence of high H2O2 concentrations possibly reflects an initial cell attempt to promote detoxification. To further demonstrate TcCPX involvement in T. cruzi response to oxidative stress, TcCPX overexpressing cells were produced. Compared to pTEX transformed cells, pTEX-TcCPX mutant cells showed a higher mRNA level (129%), without a corresponding increase in protein production (11%), suggesting that regulation of gene expression occurs at post-transcriptional levels. Furthermore, parasite treatment with 200 microM H2O2 for 30 min, led to an increase in mRNA (192%), but not in protein levels (24%). Higher mRNA levels correlated to protein levels were observed only after longer H2O2 incubation periods (1-2 h), suggesting that protein translation occurs accordingly to parasite needs. An increase in glucose-6-phosphate dehydrogenase activity was observed in pTEX-TcCPX epimastigotes that could provide cells with extra reducing power and a higher growth index.
Insights
Trypanosoma cruzi resistance to oxidative stress involves tryparedoxin peroxidase (TcCPX). TcCPX levels increase with hydrogen peroxide exposure, and its overexpression enhances parasite resistance and growth.
Area of Science:
- Parasitology
- Molecular Biology
- Biochemistry
Background:
- Trypanosoma cruzi, an intracellular parasite, faces oxidative stress from reactive oxygen species.
- Understanding hydroperoxide detoxification, including tryparedoxin peroxidase (TcCPX), is crucial for Chagas' disease chemotherapy.
- TcCPX's role in T. cruzi's defense against oxidative damage requires further investigation.
Purpose of the Study:
- To investigate the involvement of TcCPX in Trypanosoma cruzi's resistance to oxidant-mediated injury.
- To explore the regulatory mechanisms of TcCPX expression under oxidative stress.
- To assess the functional impact of TcCPX on parasite proliferation and resistance.
Main Methods:
- Exposure of T. cruzi to varying hydrogen peroxide (H2O2) concentrations.
- Western blotting to detect TcCPX protein levels.
- Generation and analysis of TcCPX-overexpressing T. cruzi cells (pTEX-TcCPX).
- Quantitative PCR (qPCR) to measure mRNA levels.
- Assay of glucose-6-phosphate dehydrogenase activity.
Main Results:
- Low H2O2 concentrations stimulated T. cruzi proliferation and increased resistance to higher H2O2 doses.
- H2O2 exposure induced a dose-dependent increase in TcCPX protein levels.
- TcCPX overexpression led to increased mRNA but not proportional protein increase, indicating post-transcriptional regulation.
- Oxidative stress primarily increased TcCPX mRNA, with protein level increases observed after prolonged exposure.
- Overexpression of TcCPX correlated with increased glucose-6-phosphate dehydrogenase activity and growth.
Conclusions:
- TcCPX plays a significant role in Trypanosoma cruzi's response to oxidative stress.
- Gene expression regulation of TcCPX occurs at post-transcriptional levels, with protein translation timed to parasite needs.
- Enhanced TcCPX activity may contribute to parasite survival and proliferation under oxidative conditions, offering potential therapeutic targets.
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